The main function of gastrointestinal (GI) smooth muscle is to mix and propel intralumenal contents, which enables efficient digestion of food, progressive absorption of nutrients, and eventual evacuation of residual components. The activity of GI smooth muscle is regulated by intrinsic and extrinsic neural signals, including classical neurotransmitters, co-existing neuropeptides, and circulating peptide hormones. In addition, a number of humoral agents including histamine, serotonin, and adenosine that are produced by nonneural GI cells also influence the activity of smooth muscle cells.
A number of clinical conditions are associated with altered GI motility, including irritable bowel syndrome, diabetic gastroparesis, postoperational ileus, chronic constipation, gastrointestinal reflux disease, chronic diarrhea, infectious diseases, malabsorptive disorders, inflammatory bowel disorders, and intestinal cancers. The identification of regulators of gastrointestinal motility should facilitate the development of novel therapeutics for disorders that involve impaired or enhanced gastrointestinal motility.
Two potential regulators of gastrointestinal motility have recently been identified. Mamba intestinal toxin (MIT1), a small protein that potently stimulates the contraction of guinea-pig ileum, has been purified from mamba snake venom (Schweitz et al., Toxicon 28:847–856 (1990) and Schweitz et al., FEBS Letters 461:183–188 (1999)). Recently, a protein of similar size and having greater than 40% identity with MIT1, including all 10 conserved cysteines, has been purified from frog skin secretions (Mollay et al., Eur. J. Pharmacol. 374:189–196 (1999)). The frog protein, named Bv8, was also found to potently stimulate the contraction of GI smooth muscle.
Methods of recombinantly preparing these snake and frog polypeptides, or of recombinantly preparing other polypeptides containing 10 cysteines, have not previously been described, limiting the utility of these regulators for therapeutic use. Additionally, snake and frog polypeptides could elicit antibodies if administered to mammals that would likely reduce their efficacy as therapeutics.
Accordingly, there exists a need to identify endogenous human polypeptides that stimulate or inhibit gastrointestinal motility, and to develop methods of preparing these compounds recombinantly as therapeutics. There also exists a need to identify small molecule agonists and antagonists of endogenous gastrointestinal regulators that can be used therapeutically. The present invention satisfies this need, and provides related advantages as well.